Nuclear hormone receptors (NRs) are key transcriptional regulators of cellular growth, differentiation, and metabolism. The NR superfamily include receptors for hormones, vitamins, metabolites, and xenobiotics, as well as receptors without known ligands, termed orphan receptors, NRs repress transcription in the absence of ligand by recruiting co-repressors N-CoR and SMRT. Hormone action is largely the result of dissociation of co-repressor, accompanied by the recruitment of co-activator molecules. A major goal of this laboratory is to understand transcriptional repression from the interaction of NRs with the co-repressors to the molecular mediators of the repressive signal. The orphan receptor RevErb has been a superb of NRs with co-repressors to the molecular mediators of the repressive signal. The orphan receptor RevErb has been a superb model for learning about activation helix, and that RevErb homodimers recruit N-CoR to specific DNA binding sites. We hypothesize that each subunit of the DNA-bound RevErb homodimer binds to a specific DNA binding sites. We hypothesize that each subunit of the DNA-bound RevErb homodimer binds to a different CoRNR motif in N-CoR. This hypothesis will be directed tested in Specific Aim 1, which will determine the molecular mechanism by which RevErb homodimers interact with co-repressor using mutagenesis, chimeric receptors and co-repressors, and proteolytic mapping. We have also identified multiple histone deacetylases (HDACs) and other proteins that interact with N-CoR/SMRT and are potential mediators of the repressive signal. We hypothesize that different co-repressors and co-repressor associated proteins are recruited by RevErb and other NRs in a receptor- and cell-specific manner. Specific Aim 2 is to determine the co-repressors and co-repressor-associated proteins involved in repression by RevErb and other NRs in vivo, using chromatin immunoprecipitation and RNA interference. We have also shown that N-CoR and SMRT act as activ ating co-factors for the HDAC3 enzyme. Specific Aim 3 is to determine the mechanism of this phenomenon, and to understand its applicability to other co-repressors and other HDACs. We hypothesize that SMRT and N-CoR have others functions related to chromatin modification and transcriptional repression. Preliminary data indicate that one such function is histone binding. Specific Aim 4 is to understand the chromatin-related function of the co-repressors. The mechanism and function of histone binding will be elucidated. Together, these studies will elucidate basic mechanisms underlying the regulation of NR interactions with co-repressors, and co-repressor regulation of transcription. The insights gained from this work will expand our understanding of the mechanisms of hormone action, and has potential to lead to novel approaches to diseases associated with NR function, including obesity, diabetes, and leukemia.